Bacterial diseases are common diseases in China's agricultural production, almost every crop has occurred, usually resulting in crop production reduction of 20% to 30%, its number and harm degree has exceeded the virus, becoming the second largest pathogen after fungus.
There are about 500 kinds of bacterial crop diseases in the world, and there are more than 200 kinds of bacterial crop diseases in China, accounting for about 1/4 to 1/3 of the types of plant bacterial diseases in the world. Rice bacterial spot, rice white leaf blight, citrus canker in fruit trees, mango canker, dragon fruit and banana soft rot, potato bacterial wilt in melons and vegetables, scab, cucumber bacterial spot, cruciferous vegetable soft rot, ginger blast, etc., all caused serious losses to crops.
Due to the characteristics of epidemic, explosive and destructive bacterial diseases, and various modes of transmission, bacterial diseases can invade from plant wounds, cracks and water holes at the edge of vegetable leaves, and can also be spread by running water, rain, insects, etc., winter in disease residues, seeds, and soil, and can easily break out in high temperature and high humidity conditions.
Crop wilt (known as "plant cancer") is one of the most devastating bacterial diseases in the world, its pathogen is Solanorella complex, can infect more than 50 families of more than 450 species of plants, global annual losses reach 1 billion US dollars. Xylella fastidiosa is a particularly dangerous plant pathogen that can infect a wide variety of crops, such as grapes, citrus, almonds, olives, peaches and coffee. The European Commission (EC) estimates that if X. fastidiosa were to be fully disseminated throughout the EU, it could result in production losses of up to €5.5 billion per year.
At present, the annual occurrence area of bacterial diseases in China is about 120 million mu. It is estimated that the current market capacity of bacterial diseases in China exceeds 3 billion yuan, mainly concentrated in the prevention and control of soilborne diseases such as orange and grapefruit ulcer, vegetable bacterial keratosis, peach tree bacterial perforation and bacterial wilt.
In recent years, the continuous use of chemical fungicides has caused a lot of plant pathogenic bacteria to produce strong resistance, and the outbreak of drug-resistant bacterial diseases is usually difficult to control, but also caused serious chemical residues and pollution, which has a great impact on agricultural production and people's lives. Bacteriophage has been widely used in medicine and food because of its advantages of strong specificity, few side effects, strong proliferation ability and not easy to produce resistance. In crops, bacteriophage has been gradually paid attention to and developed in recent years.
Basic principles of bacteriophages
Bacteriophages, a class of viruses that can specifically infect and destroy specific bacteria, have the potential to become biocontrol agents for bacterial diseases in plants. The phage binds to the bacteria through specific receptors and injects its genetic material, using the bacteria's biosynthetic system to replicate. During replication, the phage encodes specific enzymes, such as endolysins and lyases, that are able to break down the cell wall of the bacteria, causing the bacteria to lyse and release new phage particles. This process can directly kill pathogens, thus reducing the occurrence of plant diseases.
Advantages and disadvantages of bacteriophage biological control
The use of bacteriophages for biological control of bacterial diseases is a method tailored to plant infection.
Compared with traditional pesticides for crop bacterial disease control, bacteriophages have the following advantages: they are widespread in the biological world; Lower concentration has good curative effect; Easy to separate; Bacteriophages are limited self-replicating viruses. They can survive and replicate only when there is a host. In the absence of a host, bacteriophages will soon die. Phage has no toxicity to the environment and will not cause pollution to the environment, which can meet the requirements of green pollution-free agriculture advocated in modern times. Bacteriophage has strong specificity, only targeting the corresponding pathogenic bacteria, but not destroying the normal flora. While the traditional pesticide control kills pathogenic bacteria, it also destroys the beneficial flora in the soil, resulting in the imbalance of soil microorganisms and causing adverse effects on the environment. The exponential proliferation ability of phage is a significant advantage of phage therapy. Bacteria can be killed with a small amount of phage preparation, while traditional pesticides need to reach a relatively high content to achieve the purpose of sterilization. In addition, the soil pesticide residue is high and the environment is polluted. Bacteria are not easy to develop resistance to bacteriophages. On the other hand, bacteriophages can also produce appropriate mutations to adapt to the variation of host bacteria, and have the ability to mutate and overcome bacterial resistance, etc. Traditional bacterial disease control methods do not have such advantages. The development of bacteriophages requires short time, low cost and easy preservation.
As an emerging biopesticide, bacteriophage can be used as a part of the comprehensive management of plant bacterial diseases and can be used in combination with other chemical agents and biological reagents, which will have broad development prospects in the future .
The emergence of phage-resistant bacteria may become one of the major limiting factors in the use of phages to control bacterial infections. However, some researchers believe that smaller frequencies of phage resistance mutants should not hinder the use of phages as biocontrol agents. Mutations in phage receptors on the surface of bacterial cells are the most common cause of drug resistance. The mutant bacteriophages obtained from wild-type bacteriophages can be used to restore their lysing activity to bacteria. Isolate new or modified bacteriophages and use a "cocktail" of multiple bacteriophages to prevent and combat microbial resistance. Some studies have also shown that combining phage therapy with antibiotics can avoid or reduce the chance of resistance . To ensure the survival of phage mixtures during long-term storage under environmental conditions, better protective agents are required .
Application of bacteriophages in plant protection
Studies have shown that phages can effectively control a variety of plant bacterial diseases. For example, bacteriophage products for bacterial spot disease in tomatoes and peppers, fire blight in apples and pears, and soft rot in potatoes have been applied in actual agricultural production. These phage products are usually applied directly to the crop in the form of a spray, which can effectively reduce the number of pathogens and reduce the occurrence of disease.
In the early 20th century, D 'Herelle and Twort discovered bacteriophages. Soon after, research began using bacteriophages to control plant diseases. In 1924, when Mallman and Hemstreet isolated pathogenic bacteria from rotting cabbage, they found bacteriophages that inhibited the growth of pathogenic bacteria. The Thomas field trial tested Stewart corn blight and showed that by treating seeds with phages that target plant pathogens, the incidence of the disease can be reduced. However, due to the lack of knowledge about phages at the time and limited data, such studies were neglected. Nearly half a century later, Civerolo used phage treatment to reduce the severity of bacterial spots (xanthomonas) on peach seedlings by 86 to 100 percent. Over the next few years, bacteriophages were used to combat several plant diseases caused by xanthomonas, agrobacterium rhizome, Solanorella, and Phytophthora oryae. At present, bacteriophages have been studied as drugs to treat plant pathogens.
Industrialization progress
The United States Environmental Protection Agency (EPA) first approved an application for registration of a phage preparation formulated from the phages Xanthomonascompestris and P syringae in 2005.
In recent years, a number of products using phages to control plant pathogens have been developed and commercialized to treat infections in a variety of crops, including spinach, tomatoes, peppers, apples, pears, peaches, cherries, almonds, walnuts, hazelnuts, and others. Research institutions and enterprises around the world are actively exploring the application of phages in plant protection. For example, the AgriPhage family of products developed by OmniLytics has been approved by the EPA in the United States for the control of bacterial diseases in tomatoes and green peppers. The active ingredient of AgriPhage is bacteriophage, which is only effective against the target bacteria and poses very low risk to the operator, the beneficial population and the environment. In addition, AgriPhage products remain effective against bacteria that have developed resistance to antibiotics, copper preparations and other products, and can be below the maximum residue limits. In addition, research institutions in Europe and Asia are also conducting research and demonstration pilots of phages in agriculture.
In India, research to develop phage-based solutions for the agricultural sector is also intensifying, with more and more phages being isolated and demonstrated to have lytic effects against bacterial plant pathogens. Ranjan et al. isolated phage φXOF4 targeting rice moss, which was capable of cracking all rice moss strains causing rice leaf blight (BLB). The study also observed a reduced incidence of BLB in seedlings raised from phage-treated seeds. Despite the impressive efficacy of bacteriophages in controlling plant pathogens, bacteriophag-based biopesticides are not yet registered as pesticides due to strict regulations. In the EU, only a quarter of active substances and plant protection products (PPPS) are permitted for use in agriculture under current regulations. The registration procedure is complex and lengthy, as it involves four main bodies: the rapporteur Member States, the European Food Safety Authority (EFSA), the DirectorGeneral for Health and Food Safety (DG SANTE), and the Standing Committee on Plants, Animals, Food and Feed (PAFF Committee). As a result, only two companies have registered phage-based biopesticides in Europe. Enviroinvest is a Hungarian company that has developed Erwiphage® Plus for the control of fire blight in apple trees and pears. APS Biocontrol Ltd. is a Scottish company that has developed bacteriophages containing Biolyse® PB to control potato soft rot, known as pectin infection. However, to date, the European Food Safety Authority has not registered phage products as plant protection products or biopesticides.
In the United States, the Environmental Protection Agency (EPA) and the Food and Drug Administration (FDA) are involved in the regulatory framework for biopesticides. OmniLytics, Inc. is the first U.S. company to receive a phage biocontrol product registration Omnilytics, part of Phagelux, has developed four different commercial products within the Agriphage product line in the United States that have been registered as biopesticides by the U.S. Environmental Protection Agency. It is commercialized by Certis USA. Its phage-based biopesticide, AgriPhage, is used for bacterial spots on tomatoes and peppers and spots caused by Campestella and Pseudomonas syringae. In addition, more recently, the company has developed new biopesticides based on bacteriophages for the control of bacterial canker in tomatoes, fire blight in apples and pears, and citrus canker.
In Japan, XylPhi-PD is another phage-based biopesticide developed by Otsuka Pharmaceutical Company for the control of Xylella fastidiosa(leaf margin fusarium)[4,5].
Challenges faced
Although phage therapy shows great potential in plant protection, it still faces some challenges in practical application. First, the stability and persistence of phages need to be further improved to adapt to the changing natural environment. Second, the host range of the phage needs to be defined to avoid having an impact on non-target bacteria. In addition, the registration and regulation of phages is also a key factor in the promotion of applications.
One problem with phage biocontrol approaches is that the phage mixture needs to be constantly updated to lyse as many newly emerging target strains as possible. This allows the phage mixture to adapt to the relevant pathogenic strain in specific situations and avoid the development of phage resistance. However, previous EU regulations (1107/2009EC) required any changes to the composition of phage mixtures to require re-registration, time-consuming and also costly, making the US approach unfeasible in the EU. Gaps and delays in legislation allowing biocontrol agents to control bacterial plant diseases in many countries are due to the constant changes in emerging target bacterial strains, but can be overcome by screening phages adapted to new strains and timely updating. In addition, legislation relating to bacteriophage biocontrol should be flexible to allow for optimal application and implementation of biocontrol agents.
Future outlook
Global pesticides are mainly crop pesticides, and the global pesticide market size will reach $76 billion in 2022 and more than $66.7 billion in 2023. Crop protection medicines continue to dominate the global pesticide market, with a projected market share of around 88% in 2023.
There are many kinds of crop pesticide products, mainly herbicides, insecticides and fungicides. In the global crop pesticide market, herbicides accounted for 45.2%, pesticides accounted for 24.8%, fungicides accounted for 20.4%, and other pesticides accounted for 9.6%. The global bactericide market will grow at a compound annual growth rate of 4.6%, with a market value of more than $11.88 billion in 2022, and the proportion of phage products should be significantly increased in the future.